Cytokines and polypeptide hormones bind to cell-surface receptors and generate second messenger molecules, via signal transduction, that alter the metabolic direction of the cell. In addition, the receptor-ligand complexes are internalized by receptor-mediated endocytosis and localized within endosome and lysosomes in the cytoplasm of the cell. We postulated that the internalized ligand may have an intracellular role and tested this idea by directly microinjecting cytokines into the cytoplasmic compartment. Microinjection of IFNgamma induced Ia expression across the species barrier and injection of TNF` induced rapid cytotoxicity and DNA fragmentation. Recently, a role for protein kinase C (PKC) ras, and phospholipase C (PLC) has been identified in the induction of macrophage surface Ia (a biological response seen normally from IFNgamma treatment). These injection studies demonstrate a direct role for PKC, ras, and PLC in the biochemical pathways that lead to Ia expression. Injection of protein kinase A (PKA) and phospholipase A2 (PLA2) had no effect. Thus, the PKC pathway can control expression of macrophage surface Ia, possibly by regulating the genes of the major histocompatibility complex and may play many other roles in the activation of macrophages.